Plasma lipopolysaccharide elevations in cattle associated with early-stage infection by Fasciola hepatica

Fasciolosis is an endemic zoonotic parasitic disease with significant impacts on human health and both animal health and production. Early post-infection impacts on the host remain unclear. The objective of this study was to determine the changes, if any, to levels of endotoxin in cattle plasma in response to early-stage infection with Fasciola hepatica. Thirty-six (36) commercial bred cattle were experimentally infected with approximately 400 viable metacercariae. Plasma lipopolysaccharide (endotoxin) levels were examined on 24 occasions from 0 h before infection to 336 h after infection using the Limulus Amoebocyte Lysate chromogenic end point assay and compared with that of six (6) uninfected control animals. Peak lipopolysaccharide levels in infected animals were reached at 52 h after infection and returned to pre-infection levels at time 144 h after infection. Infected animals had significantly elevated lipopolysaccharide levels between 24 and 120 h after infection when compared to uninfected animals. The mean change in endotoxin units (EU)/mL over time after infection was statistically significant in infected animals. Elevations of lipopolysaccharide occurred in all infected animals suggesting a possible repeatable and titratable endotoxemia conducive to therapeutic agent model development.     

小麦饲料中添加木聚糖酶对高原型藏羊胃肠道发育的影响

旨在分析小麦粉中添加木聚糖酶(xylanase, Xyl)对藏羊羔羊胃肠道发育的影响。选择同质性良好的2～3月龄高原型藏羊公羔60只,随机分为两组,每组30只,各组内设置5个重复。对照组(Control group)饲喂不含木聚糖酶的日粮,试验组(Test group)饲喂含0.2%木聚糖酶的日粮,试验分为预饲期10 d和正试期90 d。饲养试验结束时,两组各随机选取5只羔羊进行屠宰,采集消化道组织置于多聚甲醛固定,并收集血液、瘤胃和空肠内容物。结果显示：1)与对照组相比,试验组瘤胃的角质层厚度、颗粒层厚度,瓣胃的乳头长度、乳头宽度、角质层厚度、颗粒层厚度,皱胃的黏膜厚度以及十二指肠的肌层厚度,空肠的绒毛高度、黏膜厚度,回肠的肌层厚度显著提高(P<0.05);2)对照组瘤胃和血清的LPS含量显著高于试验组(P<0.05);3)相较于对照组,试验组瘤胃的糜蛋白酶、胰蛋白酶、脂肪酶活性以及空肠的糜蛋白酶、α-淀粉酶活性显著提高(P<0.05);4)试验组瘤胃的总抗氧化能力水平、谷胱甘肽过氧化物酶及过氧化氢酶活性以及空肠的总抗氧化能力水平、谷胱甘肽过氧化物酶、超氧化物歧化酶...     

棕榈粕替代部分玉米对藏羊母羊小肠形态发育、消化酶活性及抗氧化功能的影响

试验旨在探究精料补充料中使用不同比例棕榈粕替代玉米对藏羊母羊肠道组织形态学、消化酶活性、pH、脂多糖以及抗氧化能力的影响。选取初始体重相近且健康的2～3月龄高原型藏羊母羊120只,随机分为4组,每组30只,每组6个重复,每个重复5只母羊,分别饲喂0%、15%、18%和21%水平的棕榈粕替代精料中玉米。试验期97d。结果显示：1)0%组与15%组间小肠各段的绒毛高度、绒毛宽度、隐窝深度、黏膜厚度以及绒毛高度/隐窝深度差异均不显著（P>0.05）;2)0%组空肠的α-淀粉酶、纤维素酶、脂肪酶及糜蛋白酶显著或极显著小于15%组（P<0.05或P<0.01）;3)0%组空肠的谷胱甘肽过氧化物酶、过氧化氢酶活性显著低于15%组与18%组（P<0.05）,相较于21%组差异不显著（P>0.05）;4)18%组与21%组空肠的脂多糖含量显著或极显著高于0%组（P<0.05或P<0.01）,与15%组相比差异不显著（P>0.05）。由此可见,棕榈粕可替代部分玉米饲喂藏羊母羊,推荐替代玉米的比例为15%。     

Comparison of bacterial endotoxin lipopolysaccharide concentrations in the blood,ovarian follicular fluid and uterine fluid: a clinical case of bovine metritis

We investigated the concentration of the bacterial endotoxin lipopolysaccharide (LPS) in the blood, ovarian follicular fluid and uterine fluid of a clinical case of bovine metritis. A 2-year-old lactating Holstein cow exhibited continuous fever >39.5°C for more than 2 weeks after normal calving. The cow produced a fetid, watery, red-brown uterine discharge from the vagina and was diagnosed with metritis. The LPS concentrations in plasma and uterine fluid were 0.94 and 6.34 endotoxin units (EU)/ml, respectively. One of seven follicles showed an extremely high level of LPS (12.40 EU/ml) compared to the other follicles (0.62–0.97 EU/ml). These results might suggest the presence of high concentration of LPS in follicles in cows with postpartum metritis.     

从许氏平鲉体表分离海分枝杆菌的鉴定及其脂多糖的抗原性分析

从许氏平鲉粘液、背鳍、鳃等部位分离筛选到同一菌株FZ09,经形态学观察、生理生化特性及热激蛋白65(HSP65)基因序列分析,鉴定菌株FZ09为海分枝杆菌。分别用酚-氯仿-甲醇法、乙醇-酚水法和传统热酚水法提取菌株FZ09的脂多糖(LPS),分析了LPS的组成和抗原性差异。SDS-PAGE结果表明,3种方法提取的LPS条带分布基本相同,主要条带分子量分别为14、16和23 ku,其中以乙醇-酚水法提取的LPS含量和纯度最高,热酚水法提取的LPS条带最多,抗原性最好。Western-blotting显示,海分枝杆菌抗血清主要与LPS的14 ku条带发生特异性免疫反应。高碘酸氧化免疫印迹结果表明,仅传统热酚水法提取的LPS在14 ku分子量区域有弱的阳性条带出现,其他方法提取LPS与抗血清的免疫反应呈阴性。高碘酸氧化ELISA反应显示,LPS经高碘酸氧化后与海分枝杆菌FZ09抗血清的反应活性降低,OD₄₀₅明显减弱。     

Extraction,Purification and Activity Analysis of Lipopolysaccharides from Escherichia coli Isolated from Swine

In order to obtain higher purity and high yield of LPS isolated from Escherichia coli, and evaluate its virulence,excrement of diarrhea piglet was collected as pathological samples, of which, some strains of Escherichia coli were isolated and identified. And furthermore, a strain of Escherichia coli was acquired by 16S rDNA identification. The LPS from this strain of Escherichia coli was isolated with the hot-phenol water method and purified by DNaseⅠ, RNaseA, proteinase K treatment and alcohol precipitation. The polysaccharide, protein and nucleic acid content of purified LPS were detected by phenol-sulfuric acid method, Bradford method and UV points spectrophotometric method. And its bioactivity and acute median lethal dose was determinated by tachypleus amebocyte lysate (TAL) method and acute toxicity experiment method, respectively. The results showed that a strain of highly pathogenic Escherichia coli was successfully isolated. The average yield of purified LPS was 3.74%, the proportion of the polysaccharide was 13.40%, the protein was 0.48%, and the nucleic acid was 0.06%. SDS-PAGE electrophoresis and silver stain showed that the bands were mainly concentrated in the range of 14 to 160 ku. The LAL bioactivity of the prepared LPS was 1.21×10⁷ EU/mg, LD₅₀ of the mouse abdominal cavity was 31.86 mg/kg. The results revealed that the diarrhea piglets in the farm was caused by pathogenic Escherichia coli,purified LPS from this strain of Escherichia coli had the advantages of higher purity, high yield and strong virulence, which would provide theoretical data for clinical prevention and treatment of diarrhea piglets.     

牛源多杀性巴氏杆菌主要荚膜群和脂多糖基因型两组三重PCR检测方法

为掌握国内牛源多杀性巴氏杆菌流行的主要荚膜群及脂多糖基因型,建立了检测多杀性巴氏杆菌(Pm)及其A、B荚膜群的三重PCR(PmAB-3PCR)以及检测3个脂多糖基因型的三重PCR(LPS-3PCR)方法,检验了其特异性和敏感性,用感染Pm小鼠的组织和人工污染Pm的牛鼻拭子样品进行了临床模拟检验,并对30株P m进行了PCR检测和传统血清学分型比较。结果显示:PmAB-3PCR特异性好,PmAB-3PCR和LPS-3PCR的DNA检测限分别为10~100 pg和1 ng,二者对菌液的检测限分别为200~2000 CFU和2000~20000 CFU。模拟临床样品PmAB-3PCR检测结果与细菌分离结果100%一致。PmAB-3PCR与琼扩试验的符合率为84%,二者检出的阳性率分别为88%和72%,差异不显著(P>0.05);LPS-3PCR与琼扩试验的符合率为60%,检出的阳性率分别为90%和50%,差异显著(P<0.05)。结果表明,建立的两组三重PCR方法准确高效且重复性好,可取代常规血清学方法用于国内牛巴氏杆菌病的诊断、流调和疫苗株筛选。     

Role of outer membrane proteins in immunity against murine salmonellosis--1. Antibody response to crude outer membrane proteins of Salmonella typhimurium

The humoral immune response to crude outer membrane proteins (comp) of S. typhimurium in mice has been characterized. Maximal and quicker antibody response was observed when 50 micrograms of comp was injected intraperitoneally. The comp of smooth C5 strain of S. typhimurium evoked antibody response to both lipopolysaccharide (LPS) and proteins. Absorption of these sera with LPS coated erythrocytes eliminated the antibodies to LPS completely, while the antibody level to protein was left unaltered. The comp from rough mutant (lacking O-specific chain of LPS) of S. typhimurium elicited antibodies to proteins but not to LPS. These results indicate the concomitant production of antibodies to Salmonella outer membrane proteins also. The significance of such antibodies in protection and diagnosis has been discussed.     

Antigen-nonspecific macrophage factors modulating the antibody response in vitro

Antibody response to an antigen involves the co-operation between three types of cells: macrophages, T cells and B cells. The cognate interactions between these cells play a fundamental role in the expression of a specific antibody response, but the last is modulated by antigen-nonspecific soluble factors produced either by macrophages or by T cells. Macrophages elaborate a spectrum of molecules modulating the function of lymphoid cells; among them are IL1 and prostaglandins of the E series, which are respectively enhancer and inhibitor of the antibody response in vitro. These molecules alter T cell and B cell activities through different mechanisms involving activation or inhibition of IL2 production, or alteration of cells surface antigens. However, the cellular events following the fixation of soluble factor on its receptors are not known.